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Dennis L. Stevens - One of the best experts on this subject based on the ideXlab platform.
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comparative efficacy of antibiotics in treating experimental clostridium septicum infection
International Journal of Antimicrobial Agents, 2018Co-Authors: Michael J Aldape, Amy E Bryant, Clifford R Bayer, Savannah Nicole Rice, Dennis L. StevensAbstract:ABSTRACT Clostridium septicum is a highly pathogenic microbe that causes Gas Gangrene in humans, and is the principal cause of spontaneous Gas Gangrene in patients with Gastrointestinal maladies, including adenocarcinoma of the colon. Despite modern approaches to manage C. septicum infection, morbidity and mortality remain high (>60%). At present, no objective in-vivo data exist supporting the current antibiotic treatment recommendations for C. septicum infection. Utilizing an established murine model of clostridial myonecrosis, this study investigated the efficacy of standard antibiotics for anaerobic Gram-positive soft tissue infections (penicillin, clindamycin, tetracycline and vancomycin) in treating C. septicum Gas Gangrene. Following intramuscular challenge with 1 × 106 colony-forming units of C. septicum, antibiotics were administered by intraperitoneal injection every 4 h for a total of four doses. At 30 h, all animals in all treatment groups survived the C. septicum challenge, compared with no survivors in the untreated controls (100% mortality by 10 h). However, by 60 h, mice treated with vancomycin exhibited 40% mortality, with no mortality observed in any other antibiotic treatment group. Microbroth dilution minimum inhibitory concentration analyses for three strains of C. septicum also demonstrated high susceptibility to penicillin, clindamycin and tetracycline, but considerably lower susceptibility to vancomycin. This study suggests that penicillin, clindamycin and tetracycline are suitable alternatives for the treatment of C. septicum infection in humans.
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spontaneous c septicum Gas Gangrene a literature review
Anaerobe, 2017Co-Authors: Ira Srivastava, Amy E Bryant, Michael J Aldape, Dennis L. StevensAbstract:As the infectious disease paradigm undergoes a subtle shift, unusual infections associated with malignancy and immunosuppression are being increasingly reported. Spontaneous or non-traumatic Clostridium septicum infection is one such unusual infection which has gained prominence. This article aims to understand the pathophysiology, clinical manifestations and current trends in diagnosing and treating this rare but deadly infection. To understand the multifactorial causation of this infection a review of published cases of spontaneous C. septicum Gas Gangrene was performed and a total of 94 such cases were identified. Several factors were analyzed for each case: age, infection location and underlying illness, presenting signs and symptoms, neutropenia, gross pathology of the colon, antibiotic use, surgical intervention, and survival. A known or occult malignancy was present in 71% patients and an overall mortality of 67% was observed.
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clostridial myonecrosis new insights in pathogenesis and management
Current Infectious Disease Reports, 2010Co-Authors: Amy E Bryant, Dennis L. StevensAbstract:Clostridial myonecrosis remains an important cause of human morbidity and mortality worldwide. Although traumatic Gas Gangrene can be readily diagnosed from clinical findings and widely available technologies, spontaneous Gas Gangrene is more insidious, and gynecologic infections due to Clostridium sordellii progress so rapidly that death often precedes diagnosis. In each case, extensive tissue destruction and the subsequent systemic manifestations are mediated directly and indirectly by potent bacterial exotoxins. The management triumvirate of timely diagnosis, thorough surgical removal of necrotic tissue, and treatment with antibiotics that inhibit toxin synthesis remains the gold standard of care. Yet, despite these measures, mortality remains 30% to 100% and survivors often must cope with life-altering amputations. Recent insights regarding the genetic regulation of toxin production, the molecular mechanisms of toxin-induced host cell dysfunction, and the roles of newly described toxins in pathogenesis suggest that novel prevention, diagnostic, and treatment modalities may be on the horizon for these devastating infections.
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clostridium perfringens phospholipase c induced platelet leukocyte interactions impede neutrophil diapedesis
Journal of Medical Microbiology, 2006Co-Authors: Amy E Bryant, Clifford R Bayer, Michael J Aldape, Randi J Wallace, Richard W Titball, Dennis L. StevensAbstract:Clostridium perfringens Gas Gangrene is a fulminant necrotizing infection in which inflammatory cells are notably absent from infected tissues but are often massed within adjacent vessels. It has been shown that C. perfringens phospholipase C (PLC) stimulates formation of large intravascular platelet/leukocyte complexes and that PLC-induced activation of platelet gpIIbIIIa plays a major role. In vivo, such aggregates contribute to microvascular thrombosis and ischaemic necrosis of tissue. However, the effects of adherent platelets on neutrophil diapedesis have not been established. The present work investigated (1) the contribution of platelet P-selectin (CD62P) to PLC-induced cellular complex formation and (2) the effects of platelet adhesion on neutrophil diapedesis. The effects of anti-gpIIbIIIa and anti-CD62P strategies on PLC-induced complex formation were measured by flow cytometry and followed by light microscopy. Both platelet gpIIbIIIa and CD62P contributed to the formation of platelet/leukocyte complexes. Specifically, gpIIbIIIa mediated the formation of large platelet/platelet aggregates that were tethered to the leukocyte principally via CD62P. Neutrophil diapedesis, quantified by a transendothelial cell migration assay and visualized by electron microscopy, was significantly reduced (>60 %) by the adherence of large platelet aggregates. It was concluded that the absence of a tissue inflammatory response in C. perfringens Gas Gangrene is due, in part, to impaired neutrophil mobility caused by large aggregates of adherent platelets induced by PLC. Further, an adjunctive immunotherapeutic strategy targeting both gpIIbIIIa and CD62P may improve the tissue inflammatory response, prevent vascular occlusion, maintain tissue viability, and reduce the need for radical amputation in patients with clostridial Gas Gangrene.
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immunization with the c domain of α toxin prevents lethal infection localizes tissue injury and promotes host response to challenge with clostridium perfringens
The Journal of Infectious Diseases, 2004Co-Authors: Dennis L. Stevens, Clifford R Bayer, Richard W Titball, Marie Jepson, Susan M Hayesschroer, Amy E BryantAbstract:Clostridium perfringens Gas Gangrene is characterized by rapid tissue destruction, impaired host response, and, often, death. Phospholipase C (alpha -toxin) is the virulence factor most responsible for these pathologies. The present study investigated the efficacy of active immunization with the C-terminal domain of alpha -toxin (Cpa247-370) in a murine model of Gas Gangrene. Primary end points of the study were survival, progression of infection, and tissue perfusion. Secondary end points, which were based on findings of histologic evaluation of tissues, included the extent of tissue destruction and microvascular thrombosis, as well as the magnitude of the tissue inflammatory response. Survival among C-domain-immunized animals was significantly greater than that among sham-immunized control animals. Furthermore, immunization with the C-domain localized the infection and prevented ischemia of the feet. Histopathologic findings demonstrated limited muscle necrosis, reduced microvascular thrombosis, and enhanced granulocytic influx in C-domain-immunized mice. We conclude that immunization with the C-domain of phospholipase C is a viable strategy for the prevention of morbidity and mortality associated with C. perfringens Gas Gangrene.
Amy E Bryant - One of the best experts on this subject based on the ideXlab platform.
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comparative efficacy of antibiotics in treating experimental clostridium septicum infection
International Journal of Antimicrobial Agents, 2018Co-Authors: Michael J Aldape, Amy E Bryant, Clifford R Bayer, Savannah Nicole Rice, Dennis L. StevensAbstract:ABSTRACT Clostridium septicum is a highly pathogenic microbe that causes Gas Gangrene in humans, and is the principal cause of spontaneous Gas Gangrene in patients with Gastrointestinal maladies, including adenocarcinoma of the colon. Despite modern approaches to manage C. septicum infection, morbidity and mortality remain high (>60%). At present, no objective in-vivo data exist supporting the current antibiotic treatment recommendations for C. septicum infection. Utilizing an established murine model of clostridial myonecrosis, this study investigated the efficacy of standard antibiotics for anaerobic Gram-positive soft tissue infections (penicillin, clindamycin, tetracycline and vancomycin) in treating C. septicum Gas Gangrene. Following intramuscular challenge with 1 × 106 colony-forming units of C. septicum, antibiotics were administered by intraperitoneal injection every 4 h for a total of four doses. At 30 h, all animals in all treatment groups survived the C. septicum challenge, compared with no survivors in the untreated controls (100% mortality by 10 h). However, by 60 h, mice treated with vancomycin exhibited 40% mortality, with no mortality observed in any other antibiotic treatment group. Microbroth dilution minimum inhibitory concentration analyses for three strains of C. septicum also demonstrated high susceptibility to penicillin, clindamycin and tetracycline, but considerably lower susceptibility to vancomycin. This study suggests that penicillin, clindamycin and tetracycline are suitable alternatives for the treatment of C. septicum infection in humans.
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spontaneous c septicum Gas Gangrene a literature review
Anaerobe, 2017Co-Authors: Ira Srivastava, Amy E Bryant, Michael J Aldape, Dennis L. StevensAbstract:As the infectious disease paradigm undergoes a subtle shift, unusual infections associated with malignancy and immunosuppression are being increasingly reported. Spontaneous or non-traumatic Clostridium septicum infection is one such unusual infection which has gained prominence. This article aims to understand the pathophysiology, clinical manifestations and current trends in diagnosing and treating this rare but deadly infection. To understand the multifactorial causation of this infection a review of published cases of spontaneous C. septicum Gas Gangrene was performed and a total of 94 such cases were identified. Several factors were analyzed for each case: age, infection location and underlying illness, presenting signs and symptoms, neutropenia, gross pathology of the colon, antibiotic use, surgical intervention, and survival. A known or occult malignancy was present in 71% patients and an overall mortality of 67% was observed.
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clostridial myonecrosis new insights in pathogenesis and management
Current Infectious Disease Reports, 2010Co-Authors: Amy E Bryant, Dennis L. StevensAbstract:Clostridial myonecrosis remains an important cause of human morbidity and mortality worldwide. Although traumatic Gas Gangrene can be readily diagnosed from clinical findings and widely available technologies, spontaneous Gas Gangrene is more insidious, and gynecologic infections due to Clostridium sordellii progress so rapidly that death often precedes diagnosis. In each case, extensive tissue destruction and the subsequent systemic manifestations are mediated directly and indirectly by potent bacterial exotoxins. The management triumvirate of timely diagnosis, thorough surgical removal of necrotic tissue, and treatment with antibiotics that inhibit toxin synthesis remains the gold standard of care. Yet, despite these measures, mortality remains 30% to 100% and survivors often must cope with life-altering amputations. Recent insights regarding the genetic regulation of toxin production, the molecular mechanisms of toxin-induced host cell dysfunction, and the roles of newly described toxins in pathogenesis suggest that novel prevention, diagnostic, and treatment modalities may be on the horizon for these devastating infections.
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clostridium perfringens phospholipase c induced platelet leukocyte interactions impede neutrophil diapedesis
Journal of Medical Microbiology, 2006Co-Authors: Amy E Bryant, Clifford R Bayer, Michael J Aldape, Randi J Wallace, Richard W Titball, Dennis L. StevensAbstract:Clostridium perfringens Gas Gangrene is a fulminant necrotizing infection in which inflammatory cells are notably absent from infected tissues but are often massed within adjacent vessels. It has been shown that C. perfringens phospholipase C (PLC) stimulates formation of large intravascular platelet/leukocyte complexes and that PLC-induced activation of platelet gpIIbIIIa plays a major role. In vivo, such aggregates contribute to microvascular thrombosis and ischaemic necrosis of tissue. However, the effects of adherent platelets on neutrophil diapedesis have not been established. The present work investigated (1) the contribution of platelet P-selectin (CD62P) to PLC-induced cellular complex formation and (2) the effects of platelet adhesion on neutrophil diapedesis. The effects of anti-gpIIbIIIa and anti-CD62P strategies on PLC-induced complex formation were measured by flow cytometry and followed by light microscopy. Both platelet gpIIbIIIa and CD62P contributed to the formation of platelet/leukocyte complexes. Specifically, gpIIbIIIa mediated the formation of large platelet/platelet aggregates that were tethered to the leukocyte principally via CD62P. Neutrophil diapedesis, quantified by a transendothelial cell migration assay and visualized by electron microscopy, was significantly reduced (>60 %) by the adherence of large platelet aggregates. It was concluded that the absence of a tissue inflammatory response in C. perfringens Gas Gangrene is due, in part, to impaired neutrophil mobility caused by large aggregates of adherent platelets induced by PLC. Further, an adjunctive immunotherapeutic strategy targeting both gpIIbIIIa and CD62P may improve the tissue inflammatory response, prevent vascular occlusion, maintain tissue viability, and reduce the need for radical amputation in patients with clostridial Gas Gangrene.
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immunization with the c domain of α toxin prevents lethal infection localizes tissue injury and promotes host response to challenge with clostridium perfringens
The Journal of Infectious Diseases, 2004Co-Authors: Dennis L. Stevens, Clifford R Bayer, Richard W Titball, Marie Jepson, Susan M Hayesschroer, Amy E BryantAbstract:Clostridium perfringens Gas Gangrene is characterized by rapid tissue destruction, impaired host response, and, often, death. Phospholipase C (alpha -toxin) is the virulence factor most responsible for these pathologies. The present study investigated the efficacy of active immunization with the C-terminal domain of alpha -toxin (Cpa247-370) in a murine model of Gas Gangrene. Primary end points of the study were survival, progression of infection, and tissue perfusion. Secondary end points, which were based on findings of histologic evaluation of tissues, included the extent of tissue destruction and microvascular thrombosis, as well as the magnitude of the tissue inflammatory response. Survival among C-domain-immunized animals was significantly greater than that among sham-immunized control animals. Furthermore, immunization with the C-domain localized the infection and prevented ischemia of the feet. Histopathologic findings demonstrated limited muscle necrosis, reduced microvascular thrombosis, and enhanced granulocytic influx in C-domain-immunized mice. We conclude that immunization with the C-domain of phospholipase C is a viable strategy for the prevention of morbidity and mortality associated with C. perfringens Gas Gangrene.
Julian I Rood - One of the best experts on this subject based on the ideXlab platform.
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cross complementation of clostridium perfringens plc and clostridium septicum α toxin mutants reveals plc is sufficient to mediate Gas Gangrene
Microbes and Infection, 2009Co-Authors: Catherine L Kennedy, John J Emmins, Jackie K. Cheung, Dena Lyras, Thomas J Hiscox, Julian I RoodAbstract:Clostridium perfringens and Clostridium septicum are the most common causes of clostridial myonecrosis or Gas Gangrene. Although they mediate a similar disease pathology, they elaborate functionally very different α-toxins. We used a reciprocal complementation approach to assess the contribution of the primary toxin of each species to disease and found that C. perfringens α-toxin (PLC) was able to mediate the gross pathology of myonecrosis even in a C. septicum background, although it could not induce vascular leukostasis. Conversely, while C. septicum α-toxin restored some virulence to a C. perfringens plc mutant, it was less active than in its native background.
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Synergistic effects of alpha-toxin and perfringolysin O in Clostridium perfringens-mediated Gas Gangrene.
Infection and immunity, 2001Co-Authors: Milena M Awad, Darren M Ellemor, John J Emmins, Richard L. Boyd, Julian I RoodAbstract:To examine the synergistic effects of alpha-toxin and perfringolysin O in clostridial myonecrosis, homologous recombination was used to construct an alpha-toxin deficient derivative of a perfringolysin O mutant of Clostridium perfringens. The subsequent strain was complemented with separate plasmids that carried the alpha-toxin structural gene (plc), the perfringolysin O gene (pfoA), or both toxin genes, and the resultant isogenic strains were examined in a mouse myonecrosis model. Synergistic effects were clearly observed in these experiments. Infection with the control strain, which did not produce either toxin, resulted in very minimal gross pathological changes, whereas the isogenic strain that was reconstituted for both toxins produced a pathology that was clearly more severe than when alpha-toxin alone was reconstituted. These changes were most apparent in the rapid spread of the disease, the gross pathology of the footpad and in the rate at which the mice had to be euthanatized for ethical reasons. Elimination of both alpha-toxin and perfringolysin O production removed most of the histopathological features typical of clostridial myonecrosis. These effects were restored when the mutant was complemented with the alpha-toxin structural gene, but reconstituting only perfringolysin O activity produced vastly different results, with regions of coagulative necrosis, apparently enhanced by vascular disruption, being observed. Reconstitution of both alpha-toxin and perfringolysin O activity produced histopathology most similar to that observed with the alpha-toxin reconstituted strain. The spreading of myonecrosis was very rapid in these tissues, and coagulative necrosis appeared to be restricted to the lumen of the blood vessels. The results of these virulence experiments clearly support the hypothesis that alpha-toxin and perfringolysin O have a synergistic effect in the pathology of Gas Gangrene.
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use of genetically manipulated strains of clostridium perfringens reveals that both alpha toxin and theta toxin are required for vascular leukostasis to occur in experimental Gas Gangrene
Infection and Immunity, 1999Co-Authors: Darren M Ellemor, Rebecca N Baird, Milena M Awad, Richard L. Boyd, Julian I Rood, John J EmminsAbstract:A hallmark of Gas Gangrene (clostridial myonecrosis) pathology is a paucity of leukocytes infiltrating the necrotic tissue. The cause of this paucity most likely relates to the observation of leukocyte aggregates at the border of the area of tissue necrosis, often within the microvasculature itself. Infecting mice with genetically manipulated strains of Clostridium perfringens type A (deficient in either alpha-toxin or theta-toxin production) resulted in significantly reduced leukocyte aggregation when alpha-toxin was absent and complete abrogation of leukocyte aggregation when theta-toxin was absent. Thus, both alpha-toxin and theta-toxin are necessary for the characteristic vascular leukostasis observed in clostridial myonecrosis.
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clostridial Gas Gangrene evidence that α and θ toxins differentially modulate the immune response and induce acute tissue necrosis
The Journal of Infectious Diseases, 1997Co-Authors: Dennis L. Stevens, Milena M Awad, Julian I Rood, Rodney K Tweten, Amy E BryantAbstract:The rapid extension of necrosis and an absence of polymorphonuclear leukocytes (PMNL) at the site of infection are two hallmarks of Clostridium perfringens Gas Gangrene. While both alpha and theta toxins profoundly affect PMNL function and viability in vitro, their roles in muscle destruction and impairment of the inflammatory response in vivo have not been investigated. Comparative histopathologic examinations were performed on animals infected with either wild-type C. perfringens, or isogenic, toxin-deficient mutants of C. perfringens. Tissue destruction was modest in animals infected with the alpha toxin-deficient mutant; destruction was more pronounced in tissues infected with the theta toxin-deficient mutant or the wild-type strain. alpha and theta toxins also displayed differing abilities to modulate the inflammatory response. Histopathologic studies in which recombinant toxins were injected together with killed, washed C. perfringens further substantiated these tissue-destructive and differential antiinflammatory effects.
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virulence studies on chromosomal alpha toxin and theta toxin mutants constructed by allelic exchange provide genetic evidence for the essential role of alpha toxin in clostridium perfringens mediated Gas Gangrene
Molecular Microbiology, 1995Co-Authors: Miilena M Awad, Amy E Bryant, Dennis L. Stevens, Julian I RoodAbstract:The pathogenesis of clostridial myonecrosis, or Gas Gangrene, involves the growth of the anaerobic bacterium Clostridium perfringens in the infected tissues and the elaboration of numerous extracellular toxins and enzymes. The precise role of each of these toxins in tissue invasion and necrosis has not been determined. To enable genetic approaches to be used to study C. perfringens pathogenesis we developed an allelic exchange method which involved the transformation of C. perfringens cells with a suicide plasmid carrying a gene insertionally inactivated with an erythromycin-resistance determinant. The frequency with which double reciprocal crossover events were observed was increased to a workable level by increasing the amount of homologous DNA located on either side of the inactivated gene. Allelic exchange was used to isolate mutations in the chromosomal pfoA gene, which encodes an oxygen-labile haemolysin known as theta-toxin or perfringolysin O, and in the chromosomal plc gene, which encodes the alpha-toxin or phospholipase C. The resultant mutants failed to produce detectable theta-toxin or alpha-toxin activity, respectively, and could be complemented by recombinant plasmids that carried the respective wild-type genes. The resultant strains were virulence tested in a mouse myonecrosis model. The results showed that the plc mutants had demonstrably reduced virulence and therefore provided definitive genetic evidence for the essential role of alpha-toxin in Gas Gangrene or clostridial myonecrosis.
Richard W Titball - One of the best experts on this subject based on the ideXlab platform.
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reactive oxygen species and the mek erk pathway are involved in the toxicity of clostridium perfringens α toxin a prototype bacterial phospholipase c
The Journal of Infectious Diseases, 2012Co-Authors: Laura Monturiolgross, Richard W Titball, Marietta Floresdiaz, Graeme C Clark, Cindy Arayacastillo, Mariajose Pinedapadilla, Alberto AlapegironAbstract:Clostridium perfringens, the most broadly distributed pathogen in nature, produces a prototype phospholipase C, also called α-toxin, which plays a key role in the pathogenesis of Gas Gangrene. α-Toxin causes plasma membrane disruption at high concentrations, but the role of intracellular mediators in its toxicity at low concentrations is unknown. This work demonstrates that α-toxin causes oxidative stress and activates the MEK/ERK pathway in cultured cells and furthermore provides compelling evidence that O(2)(-.), hydrogen peroxide, and the OH(.) radical are involved in its cytotoxic and myotoxic effects. The data show that antioxidants and MEK1 inhibitors reduce the cytotoxic and myotoxic effects of α-toxin and demonstrate that edaravone, a clinically used hydroxyl radical trap, reduces the myonecrosis and the mortality caused by an experimental infection with C. perfringens in a murine model of Gas Gangrene. This knowledge provides new insights for the development of novel therapies to reduce tissue damage during clostridial myonecrosis.
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clostridium perfringens phospholipase c induced platelet leukocyte interactions impede neutrophil diapedesis
Journal of Medical Microbiology, 2006Co-Authors: Amy E Bryant, Clifford R Bayer, Michael J Aldape, Randi J Wallace, Richard W Titball, Dennis L. StevensAbstract:Clostridium perfringens Gas Gangrene is a fulminant necrotizing infection in which inflammatory cells are notably absent from infected tissues but are often massed within adjacent vessels. It has been shown that C. perfringens phospholipase C (PLC) stimulates formation of large intravascular platelet/leukocyte complexes and that PLC-induced activation of platelet gpIIbIIIa plays a major role. In vivo, such aggregates contribute to microvascular thrombosis and ischaemic necrosis of tissue. However, the effects of adherent platelets on neutrophil diapedesis have not been established. The present work investigated (1) the contribution of platelet P-selectin (CD62P) to PLC-induced cellular complex formation and (2) the effects of platelet adhesion on neutrophil diapedesis. The effects of anti-gpIIbIIIa and anti-CD62P strategies on PLC-induced complex formation were measured by flow cytometry and followed by light microscopy. Both platelet gpIIbIIIa and CD62P contributed to the formation of platelet/leukocyte complexes. Specifically, gpIIbIIIa mediated the formation of large platelet/platelet aggregates that were tethered to the leukocyte principally via CD62P. Neutrophil diapedesis, quantified by a transendothelial cell migration assay and visualized by electron microscopy, was significantly reduced (>60 %) by the adherence of large platelet aggregates. It was concluded that the absence of a tissue inflammatory response in C. perfringens Gas Gangrene is due, in part, to impaired neutrophil mobility caused by large aggregates of adherent platelets induced by PLC. Further, an adjunctive immunotherapeutic strategy targeting both gpIIbIIIa and CD62P may improve the tissue inflammatory response, prevent vascular occlusion, maintain tissue viability, and reduce the need for radical amputation in patients with clostridial Gas Gangrene.
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immunization with the c domain of α toxin prevents lethal infection localizes tissue injury and promotes host response to challenge with clostridium perfringens
The Journal of Infectious Diseases, 2004Co-Authors: Dennis L. Stevens, Clifford R Bayer, Richard W Titball, Marie Jepson, Susan M Hayesschroer, Amy E BryantAbstract:Clostridium perfringens Gas Gangrene is characterized by rapid tissue destruction, impaired host response, and, often, death. Phospholipase C (alpha -toxin) is the virulence factor most responsible for these pathologies. The present study investigated the efficacy of active immunization with the C-terminal domain of alpha -toxin (Cpa247-370) in a murine model of Gas Gangrene. Primary end points of the study were survival, progression of infection, and tissue perfusion. Secondary end points, which were based on findings of histologic evaluation of tissues, included the extent of tissue destruction and microvascular thrombosis, as well as the magnitude of the tissue inflammatory response. Survival among C-domain-immunized animals was significantly greater than that among sham-immunized control animals. Furthermore, immunization with the C-domain localized the infection and prevented ischemia of the feet. Histopathologic findings demonstrated limited muscle necrosis, reduced microvascular thrombosis, and enhanced granulocytic influx in C-domain-immunized mice. We conclude that immunization with the C-domain of phospholipase C is a viable strategy for the prevention of morbidity and mortality associated with C. perfringens Gas Gangrene.
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identification of residues critical for toxicity in clostridium perfringens phospholipase c the key toxin in Gas Gangrene
FEBS Journal, 2000Co-Authors: Richard W Titball, Bruno Lomonte, Marietta Floresdiaz, Alberto Alapegiron, Isabelle Guillouard, Claire E Naylor, Alexandra Rucavado, Ajit K Basak, Jose Maria GutierrezAbstract:Clostridium perfringens phospholipase C (PLC), also called alpha-toxin, is the major virulence factor in the pathogenesis of Gas Gangrene. The toxic activities of genetically engineered alpha-toxin variants harboring single amino-acid substitutions in three loops of its C-terminal domain were studied. The substitutions were made in aspartic acid residues which bind calcium, and tyrosine residues of the putative membrane-interacting region. The variants D269N and D336N had less than 20% of the hemolytic activity and displayed a cytotoxic potency 103-fold lower than that of the wild-type toxin. The variants in which Tyr275, Tyr307, and Tyr331 were substituted by Asn, Phe, or Leu had 11-73% of the hemolytic activity and exhibited a cytotoxic potency 102- to 105-fold lower than that of the wild-type toxin. The results demonstrated that the sphingomyelinase activity and the C-terminal domain are required for myotoxicity in vivo and that the variants D269N, D336N, Y275N, Y307F, and Y331L had less than 12% of the myotoxic activity displayed by the wild-type toxin. This work therefore identifies residues critical for the toxic activities of C. perfringens PLC and provides new insights toward understanding the mechanism of action of this toxin at a molecular level.
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structure of the key toxin in Gas Gangrene
Nature Structural & Molecular Biology, 1998Co-Authors: Claire E Naylor, Richard W Titball, J T Eaton, Angela M Howells, Neil Justin, D S Moss, Ajit K BasakAbstract:Clostridium perfringens α-toxin is the key virulence determinant in Gas Gangrene and has also been implicated in the pathogenesis of sudden death syndrome in young animals. The toxin is a 370-residue, zinc metalloenzyme that has phospholipase C activity, and can bind to membranes in the presence of calcium. The crystal structure of the enzyme reveals a two-domain protein. The N-terminal domain shows an anticipated structural similarity to Bacillus cereus phosphatidylcholine-specific phospholipase C (PC-PLC). The C-terminal domain shows a strong structural analogy to eukaryotic calcium-binding C2 domains. We believe this is the first example of such a domain in prokaryotes. This type of domain has been found to act as a phospholipid and/or calcium-binding domain in intracellular second messenger proteins and, interestingly, these pathways are perturbed in cells treated with α-toxin. Finally, a possible mechanism for α-toxin attack on membrane-packed phospholipid is described, which rationalizes its toxicity when compared to other, non-haemolytic, but homologous phospholipases C.
Alberto Alapegiron - One of the best experts on this subject based on the ideXlab platform.
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reactive oxygen species and the mek erk pathway are involved in the toxicity of clostridium perfringens α toxin a prototype bacterial phospholipase c
The Journal of Infectious Diseases, 2012Co-Authors: Laura Monturiolgross, Richard W Titball, Marietta Floresdiaz, Graeme C Clark, Cindy Arayacastillo, Mariajose Pinedapadilla, Alberto AlapegironAbstract:Clostridium perfringens, the most broadly distributed pathogen in nature, produces a prototype phospholipase C, also called α-toxin, which plays a key role in the pathogenesis of Gas Gangrene. α-Toxin causes plasma membrane disruption at high concentrations, but the role of intracellular mediators in its toxicity at low concentrations is unknown. This work demonstrates that α-toxin causes oxidative stress and activates the MEK/ERK pathway in cultured cells and furthermore provides compelling evidence that O(2)(-.), hydrogen peroxide, and the OH(.) radical are involved in its cytotoxic and myotoxic effects. The data show that antioxidants and MEK1 inhibitors reduce the cytotoxic and myotoxic effects of α-toxin and demonstrate that edaravone, a clinically used hydroxyl radical trap, reduces the myonecrosis and the mortality caused by an experimental infection with C. perfringens in a murine model of Gas Gangrene. This knowledge provides new insights for the development of novel therapies to reduce tissue damage during clostridial myonecrosis.
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role of clostridium perfringens phospholipase c in the pathogenesis of Gas Gangrene
Toxicon, 2003Co-Authors: Marietta Floresdiaz, Alberto AlapegironAbstract:Gas Gangrene is an acute and devastating infection most frequently caused by Clostridium perfringens and characterized by severe myonecrosis, intravascular leukocyte accumulation, and significant thrombosis. Several lines of evidence indicate that C. perfringens phospholipase C (Cp-PLC), also called alpha-toxin, is the major virulence factor in this disease. This toxin is a Zn2+ metalloenzyme with lecithinase and sphingomyelinase activities. Its three dimensional structure shows two domains, an N-terminal domain which contains the active site, and a C-terminal domain required for the Ca2+dependent interaction with membranes. Cp-PLC displays several biological activities: it increases capillary permeability, induces platelet aggregation, hemolysis, myonecrosis, decreases cardiac contractility, and is lethal. Experiments with genetically engineered Cp-PLC variants have revealed that the sphingomyelinase activity and the C-terminal domain are required for toxicity. The myotoxicity of Cp-PLC is largely dependent on its membrane damaging effect. In addition, it has been suggested that the alterations in the blood flow induced by this toxin also contribute to muscle damage. In Gas Gangrene, Cp-PLC dysregulates transduction pathways in endothelial cells, platelets and neutrophils leading to the uncontrolled production of several intercellular mediators and adhesion molecules. Thus, Cp-PLC alters the traffic of neutrophils to the infected tissue and promotes thrombotic events, enhancing the conditions for anaerobic growth.
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identification of residues critical for toxicity in clostridium perfringens phospholipase c the key toxin in Gas Gangrene
FEBS Journal, 2000Co-Authors: Richard W Titball, Bruno Lomonte, Marietta Floresdiaz, Alberto Alapegiron, Isabelle Guillouard, Claire E Naylor, Alexandra Rucavado, Ajit K Basak, Jose Maria GutierrezAbstract:Clostridium perfringens phospholipase C (PLC), also called alpha-toxin, is the major virulence factor in the pathogenesis of Gas Gangrene. The toxic activities of genetically engineered alpha-toxin variants harboring single amino-acid substitutions in three loops of its C-terminal domain were studied. The substitutions were made in aspartic acid residues which bind calcium, and tyrosine residues of the putative membrane-interacting region. The variants D269N and D336N had less than 20% of the hemolytic activity and displayed a cytotoxic potency 103-fold lower than that of the wild-type toxin. The variants in which Tyr275, Tyr307, and Tyr331 were substituted by Asn, Phe, or Leu had 11-73% of the hemolytic activity and exhibited a cytotoxic potency 102- to 105-fold lower than that of the wild-type toxin. The results demonstrated that the sphingomyelinase activity and the C-terminal domain are required for myotoxicity in vivo and that the variants D269N, D336N, Y275N, Y307F, and Y331L had less than 12% of the myotoxic activity displayed by the wild-type toxin. This work therefore identifies residues critical for the toxic activities of C. perfringens PLC and provides new insights toward understanding the mechanism of action of this toxin at a molecular level.
